A novel low-molecular weight chondroitin sulphate proteoglycan isolated from cartilage.
Proteoglycans were isolated from cartilage by extraction with 4M-guanidinium chloride followed by direct centrifugation in 4M-guanidinium chloride/CsCl at a low starting density, 1.34 g/ml. N-Ethylmaleimide was included in the extraction solvent as a precaution against contamination of proteoglycans with unrelated proteins mediated by disulphide exchange. A novel, discrete, low-buoyant-density proteoglycan (1.40--1.35 g/ml) was demonstrated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. Its proteoglycan nature was revealed by the shift in the molecular size observed on gel electrophoresis after treatment with chondroitinase ABC. The core protein was monodisperse. The proteoglycan was further purified by gel chromatography with and without addition of hyaluronate. The proteoglycan constitutes less than 2% (by weight) of the total extracted proteoglycans and is not capable of interacting with hyaluronate. The same proteoglycan was purified in larger quantities by sequential associative and dissociative CsCl-density-gradient centrifugation, zonal rate sedimentation in a sucrose gradient and gel chromatography on Sepharose CL-4B. The pure proteoglycan had a molecular weight of 76 300 determined by sedimentation-equilibrium centrifugation and an apparent partial specific volume of 0.59 ml/g. It contained about 25% protein (of dry weight) and had remarkably high contents of leucine and cysteine as compared with other proteoglycans. The proteoglycan contained two to three large chondroitin sulphate chains and some oligosaccharides.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D002356:Cartilage / Q000032:analysis*; D002417:Cattle; D002499:Centrifugation, Density Gradient; D002807:Chondroitin / Q000031:analogs & derivatives*; D011508:Chondroitin Sulfate Proteoglycans / Q000302:isolation & purification*; D002809:Chondroitin Sulfates / Q000032:analysis*; D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D008970:Molecular Weight; D011489:Protein Denaturation; D011509:Proteoglycans / Q000302:isolation & purification*
D000596:Amino Acids; D002241:Carbohydrates; D011508:Chondroitin Sulfate Proteoglycans; D011509:Proteoglycans; D002807:Chondroitin; D002809:Chondroitin Sulfates